Field of the Invention
In one of its aspects, the present invention relates to a method for quantifying the ultraviolet (UV) fluence received by a fluid. In another of its aspects, the present invention relates to a method for UV treatment of a fluid containing a fluorescent composition of matter. In another of its aspects, the present invention relates to a method for UV treatment of cell culture media. In another of its aspects, the present invention relates to a method for UV treatment of culture media used in the production of biopharmaceuticals. In another of its aspects, the present invention relates to a method for UV treatment of fluids used in the purification of biopharmaceuticals. In another of its aspects, the present invention relates to a system for determining the UV fluence received by a fluid being treated in a UV fluid treatment system.
Description of the Prior Art
Ultraviolet (UV) radiation is commonly used to disinfect many types of fluid media by inactivating microorganisms such as bacteria, protozoa and viruses that may be present in those fluids. This type of sterilization is favorable as a non-thermal and non-adulterating process and is used in various industries and applications, including in the biopharmaceutical industry.
While UV irradiation has been used in the biopharmaceutical industry for packaging and surface sterilization applications, its application to cell culture media as a sterilization method has been very limited elsewhere. Production growth media or culture media are liquids comprising complex mixtures of amino acids, sugar, vitamins and other compounds designed to support the growth of microorganisms or cells.
Determining the UV fluence received by a fluid is important to ensure that an effective dose (also referred to throughout this specification as “fluence”) of UV radiation has been received by the fluid to effectively inactivate the microorganisms present in the fluid. If too little UV radiation is received by the fluid, microorganisms in the fluid will not be inactivated to the extent required. Alternatively, certain fluid media may not tolerate over-irradiation and may be damaged if the UV fluence is not relatively precise.
Generally, UV radiation is applied to a fluid via UV emitters (e.g., UV lamps and the like) provided in a fluid treatment zone of a flow-through reactor. The UV fluence applied to a fluid in such a flow-through reactor is a function of, for example, the reactor design, lamp output, flow rate, as well as properties of the fluid itself (such as turbidity or opacity).
There are several known methods to measure the UV fluence in such flow-through reactors. One method consists of monitoring the UV source output (lamp intensity), optical absorbance and flow rate of the fluid to estimate UV fluence. This method has the disadvantage of being an indirect measure of UV fluence received by the fluid, and also is one which is not capable of accounting for non-uniform UV source output, blocked or non-uniform flow paths within the reactor, and is further dependent on accurate flow rate measurements.
UV actinometry is a known method to quantify the amount of UV radiation applied to a fluid. According to typical actinometric techniques, an exogenous UV-sensitive compound with a known quantum yield is added to the fluid at a concentration sufficient to absorb all incident photons. The UV-sensitive compound undergoes a UV-induced chemical change. The concentration of the photo-product produced by application of UV radiation to the UV sensitive compound is then measured. Together with the known quantum yield, the photo-product concentration can be used to quantify the absorbed UV radiation. Common, known UV actinometers include the ferrioxylate actinometer and the iodide/iodate actinometer.
Such exogenous actinometry solutions can be passed through a UV reactor before or after treatment of the target fluid in order to determine the UV fluence before or after the treatment, but cannot determine the UV fluence during treatment without changing the fluid composition.
U.S. Pat. No. 7,993,580 [Anderle et al. (Anderle)] purportedly addresses this limitation of conventional exogenous actinometric techiniques through the use of a separate flow path with a thin layer of actinometry solution in a UV reactor, and using the measured change in the chemistry of the actinometer to control the reactor. However, this method requires a separate flow path for the actinometry solution to prevent contamination of the process fluid, and therefore does not measure the dose actually delivered to the process fluid. The separate flow path of the method taught by Anderle introduces complexity and uncertainty by both requiring a separate flow path and by not directly measuring the UV fluence applied to the process fluid itself.
International Publication Number WO 2003/007998 [Li et al.] teaches a method of monitoring UV irradiation of a fluid containing protein through changes in optical absorbance at 314 nm. This method has limited sensitivity. Absorbance measurements suffer from interference from other compounds. Many constituents in growth media will contribute to the absorbance at 314 nm (or any other wavelength). Therefore, there is an increased likelihood that changes in absorbance of one compound will be masked by absorbance of other compounds that do not change with UV dose.
The photochemistry of pure actinometry methods can be well characterized, but such methods become considerably more complicated when the fluid is a complex mixture of organic and inorganic molecules, such as a cell culture medium. In such complex fluids, chemical reactions induced by absorption of UV radiation by other species can result in other reactions with the actinometric compound in addition to the desired reaction of the actinometric compound to produce the actinometric photo-product, thereby interfering with calculation of the concentration of the photo-product and in turn the UV fluence received by the fluid.
Moreover, the addition of actinometric compounds to process fluids may not be permitted in certain applications. For example, the chemical composition of biopharmaceutical growth media (so-called “upstream” fluids) and biopharmaceutical production fluids (so-called “downstream” fluids) are tightly controlled and subject to rigorous validation and regulatory approval. For this reason, exogenous actinometers are typically not suitable to monitor UV fluence in fluids used in the course of biopharmaceutical production and purification.
There is therefore a need for an endogenous actinometric process by which the UV fluence delivered to a complex fluid (e.g., a cell culture medium) can be measured to ensure that such fluids have achieved a target level of disinfection.